Examples
Table I: Typical acceptance criteria.
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As noted, assessing homogeneity for preclinical dose formulations is typically assessed by analyzing samples taken from the
top, middle, and bottom of a formulation batch using a validated analytical method. Two samples are removed from each stratum,
for analysis resulting in a sampling size of n=6. The sample results are compared with standard acceptance criteria for precision
(percent relative standard deviation or % RSD). Typical acceptance criteria are featured in Table I.
Table II: Homogeneity results. RSD is relative standard deviation
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On rare occasions, homogeneity assessments fail and it is necessary to modify the procedure for the preparation of a dosing
formulation. For example, a feed study being dosed at 1 mg/g produced the following homogeneity results: The variability observed
in Table II indicated the possibility that the material was not sufficiently mixed or the test article was not uniform. Homogeneity
was improved significantly by grinding the test article in a pestle with a mortar, followed by sieving the material to ensure
uniformity prior its addition to feed.
A second example of a failed homogeneity assessment involved a suspension. A test batch was prepared prior to the study to
establish an appropriate preparation procedure. Analysis of the test batch indicated that the material was homogeneous. However,
analysis of the initial study batch indicated the dose formulation was not homogeneous. Observed differences between the test
and study batch were batch size, mixing techniques (related to batch size), and lot of test article.
Following an extensive investigation, it was determined that the lot of test article adversely affected homogeneity. The test
articles used in the preparations were sourced from two different manufacturers; the one used for the study batch was not
of uniform particle size. To prepare a homogenous dose formulation using the second lot of material required grinding with
a mortar in a pestle and then using a sieve to ensure uniform particle size.
In another example, a dose formulation suspension prepared in 0.5% aqueous methylcellulose, required several modifications
prior to successfully preparing a homogenous dosing formulation. In this instance, multiple test batches were prepared using
various homogenization and sonication times. The final preparation required dry grinding of the test article using a pestle
and mortar. The material was subjected to additional grinding after the addition of small volumes of vehicle to form a smooth
paste. The remaining volume of vehicle was added and the formulation was mixed for 5 min using a Silverson homogenizer, sonicated
for 30 min and then stirred for 1 h. Subjecting the formulation to extensive mixing and sonication were required to prepare
a homogenous material.
Conclusion
The preparation of a homogeneous dosing formulation is a critical component of any preclinical study. Homogeneity depends
upon various factors, including batch size, mixing and blending techniques, and particle size. Accurately determining homogeneity
depends upon standardized sampling techniques and using appropriate analytical methodology.
If possible, the use of test batches prior to study initiation can go far in predetermining homogeneity. If the preparation
methodology is appropriate, it can facilitate successful study execution.
Amy Smith is director, and Melissa Whitsel is analytical manager, both with MPI Research; headquarters located at 54943 North Main Street, Mattawan, MI 49071.
References
1. FDA 21 CFR Part 58, Good Laboratory Practice Regulations, Final Rule.
2. M. Whitmire et al., AAPS Jrnl.,
12 (4), 628–634 (2010).
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